Structural studies of the C-terminal 19-peptide of serum amyloid A and its Pro→Ala variants interacting with human cystatin C

Abstract

Serum amyloid A (SAA) is a multifunctional acute-phase protein whose concentration in serum increases markedly following a number of chronic inflammatory and neoplastic diseases. Prolonged high SAA level may give rise to reactive systemic amyloid A (AA) amyloidosis, where the N-terminal segment of SAA is deposited as amyloid fibrils. Besides, recently, well-documented association of SAA with high-density lipoprotein or glycosaminoglycans, in particular heparin/heparin sulfate (HS), and specific interaction between SAA and human cystatin C (hCC), the ubiquitous inhibitor of cysteine proteases, was proved. Using a combination of selective proteolytic excision and high-resolution mass spectrometry, a hCC binding site in the SAA sequence was determined as SAA(86-104). The role of this SAA C-terminal fragment as a ligand-binding locus is still not clear. It was postulated important in native SAA folding and in pathogenesis of AA amyloidosis. In the search of conformational details of this SAA fragment, we did its structure and affinity studies, including its selected double/triple Pro → Ala variants. Our results clearly show that the SAA(86-104) 19-peptide has rather unordered structure with bends in its C-terminal part, which is consistent with the previous results relating to the whole protein. The results of affinity chromatography, fluorescent ELISA-like test, CD and NMR studies point to an importance of proline residues on structure of SAA(86-104). Conformational details of SAA fragment, responsible for hCC binding, may help to understand the objective of hCC-SAA complex formation and its importance for pathogenesis of reactive amyloid A amyloidosis.

Method development and characterization of the low molecular weight peptidome of human wound fluids

“…SAA1 can upregulate inflammatory cytokines, whereas SAA1 peptides suppress inflammation 58 . Moreover, the C-terminal SAA(86-104) binds and inhibits human cystatin and carboxyterminal fragments are involved in amyloid diseases 59 . Interestingly, similar C-terminal fragments were found in non-infected wounds, indicating that SAA is proteolyzed in wounds, suggesting a yet undisclosed physiological role for such fragments during normal wound healing.…”

Structure and Expression of Different Serum Amyloid A (SAA) Variants and their Concentration-Dependent Functions During Host Insults

“…Likewise, SAA1 and SAA2 have an amino acid sequence [tyrosine (Y) – isoleucine (I) – glycine (G) – serine (S) – aspartic acid (D)] between residues 29 and 33, comparable to that of the cell binding site of laminin [tyrosine (Y) – isoleucine (I) – glycine (G) – serine (S) – arginine (R)], which is also an extracellular matrix protein. Recently, amino acids 86-104 and particularly the proline (P) residues were found to be important for binding of cystatin C to A-SAA [38]. No data are yet available about receptor-binding sites in the SAA proteins.…”

Comparative Study on Hyaluronic Acid Binding to Murine SAA1.1 and SAA2.2

“…49 Although the transition of the SAA dimeric interface happens rapidly during the first 100 ns of the simulation, the configuration of the C-terminal loop is preserved along with the simulation, which might be due to the relatively low B-factor of this region in the crystal structure and in accordance with in vitro findings. 3,49 Functions of the SAA C-terminus include GAGs binding as well as modulation of the SAA fibril formation. 2,50 Still, the mechanism of the C-terminus-mediated fibril formation is ambiguous.…”